Successful wound healing requires the coordinated activities of the multiple cell types that constitute the inflammatory and reparatory response to tissue injury. The identification of growth factors, cytokines, extracellular matrix components and other compounds present in the wound promises clinical applications that will allow active therapeutic intervention during the healing process. The rational development of these applications will require, in turn, a better understanding of the cell biology of the inflammatory cells, most specifically as it relates to its modulation by conditions unique to the wound environment. It is the basis of this proposal that the regulation of inflammatory cell function cannot be fully understood unless the peculiarities of the environment in which these cells live and receive and react to specific signals is taken into consideration. In this regard, work in this laboratory demonstrated that a prominent feature of healing wounds is a decreased arginine concentration that results from the metabolism of this amino acid through the inducible form of nitric oxide synthase (iNOS) and arginase, and that these two enzymes are expressed in the wound in a temporally segregated manner. iNOS is expressed prominently during the very early phases of inflammation and arginase is the only detectable high-flux enzyme of a?ginine metabolism in the wound at the time of predominant macrophage/fibroblast infiltration. iNOS expression at-that time is actively suppressed by conditions prevailing in the wound, including the aforementioned reduced arginine concentration, hypoxia and by molecules present in wound fluid. It is the central hypothesis o this proposal that iNOS is a "forbidden pathway" in healing wounds because its products suppress cell functions relevant to repair (i.e.: macrophage phagocytosis, fibroblast proliferation and collagen deposition) and ultimately mediate cell death. To test this hypothesis, the Specific Aims of this prpposal are designed to define the functional phenotype of macrophages obtained from experimental wounds in the rat; evaluate the impact of arginine availability, wound fluids and extracellular matrix components in the acquisition and maintenance of this phenotype; investigate the role of arginine and NO in the regulation of macrophage/fibroblast interactions; and define the mechanisms of metabolic injury associated with the production of NO. Additionally, work proposed here will explore the role of apoptosis and the removal of apoptotic cells from the wound by wound macrophages in the termination of inflammation and repair.